Method of promoting mucosal hydration with certain uridine, adenine and cytidine diphosphates and analogs thereof

ABSTRACT

A method and preparation for the stimulation of mucosal hydration in a subject in need of such treatment is disclosed. The method comprises administering to the mucosal surfaces of the subject a purinergic receptor agonist such as uridine 5′-diphosphate (UDP), dinucleotide triphosphates; cytidine 5′-diphosphate (CDP), adenosine 5′-diphosphate (ADP), or their therapeutically useful analogs and derivatives, in an amount effective to stimulate mucin secretion. Pharmaceutical formulations and methods of making the same are also disclosed. Methods of administering the same would include: topical administration via a liquid, gel, cream, or as part of a contact lens or selective release membrane; or systemic administration via nasal drops or spray, inhalation by nebulizer or other device, oral form (liquid or pill), injectable, intra-operative instillation or suppository form. A method for facilitating the expectoration of sputum for the purpose of detecting cellular abnormalities indicative of lung disease is also disclosed.

[0001] This application is a continuation of U.S. application Ser. No.09/512,867, filed Feb. 25, 2000; which claims the benefit of U.S.Provisional Application No. 60/121,754, filed Feb. 26, 1999.

TECHNICAL FIELD

[0002] This invention relates to a method of regulating secretions inand around the mucous membranes of a mammal by administering purinergicreceptor agonists such as certain uridine, adenine, or cytidinediphosphates as well as other nucleoside phosphate compounds.

BACKGROUND OF THE INVENTION

[0003] There are many situations where it is therapeutically desirableto increase the amount of hydration on mucosal surfaces of the body.Mucus membranes are hydrated surface epithelial tissues that linecavities exposed to the outside environment. These mucosal surfaces ofthe body must stay hydrated with the proper mixtures of water, salt,mucin and other proteins in order to defend the body from the outsideworld and remain comfortable. The mucus membranes play a major role infighting off infections and keeping the exposed organs clean andhealthy. Mucus membranes are found on the surface of the eye, vagina,sinonasal cavities and mouth (oropharyngeal). Impaired hydration ofthese mucus membranes leads to medical conditions such as: dry eye,vaginal dryness, rhinosinusitis, dry mouth, corneal injury, and others.

[0004] Dry eye disease is the general term for indications in which themucous membranes surrounding the eye are not properly hydrated due to animbalance of salt, water and mucus in the precorneal tear film. Dry eyeis characterized by a decrease in tear production or an increase in tearfilm evaporation, together with the ocular surface disease that results.Approximately 38 million Americans are affected with some type of dryeye disorder. Among the indications that are referred to by the generalterm “dry eye disease” are: keratoconjunctivitis sicca (KCS),age-related dry eye, Stevens-Johnson syndrome, Sjogren's syndrome,ocular cicatrical pemphigoid, blepharitis, corneal injury, infection,Riley-Day syndrome, congenital alacrima, nutritional disorders ordeficiencies (including vitamin deficiencies), pharmacologic sideeffects, eye stress and glandular and tissue destruction, environmentalexposure to smog, smoke, excessively dry air, airborne particulates,autoimmune and other immunodeficient disorders, and comatose patientsrendered unable to blink. The present invention may also be useful as awash or irrigation solution in conscious individuals, during surgery orto maintain comatose patients or those who cannot blink due toneuromuscular blockade or loss of the eyelids.

[0005] Currently, the pharmaceutical treatment of dry eye disease ismostly limited to administration of artificial tears (saline solution)to temporarily rehydrate the eyes. However, relief is short-lived andfrequent dosing is necessary. In addition, artificial tears often havecontra-indications and incompatibility with soft contact lenses (M.Lemp, Cornea 9(1):S48-550 (1990)). The use of phosphodiesteraseinhibitors, such as 3-isobutyl-1-methylxanthine (IBMX) to stimulate tearsecretion is disclosed in U.S. Pat. No. 4,753,945. The effectiveness ofthese phosphodiesterase inhibitors is currently being investigated (J.Gilbard, et al., Arch. Ophthal. 112:1614-16 (1994) and 109:672-76(1991); idem, Inv. Ophthal. Vis. Sci. 31:1381-88 (1990)). Stimulation oftear secretion by topical application of melanocyte stimulating hormonesis described in U.S. Pat. No. 4,868,154.

[0006] Vaginal dryness is a very common problem which brings physicaland emotional distress to many women (E. Key, Nurs. Stand. 5:24-27(1991)). It most commonly manifests itself during sexual intercourse,which causes dyspareunia and can eventually lead to apareunia. Althoughit is traditionally considered to be a condition which affectspostmenopausal women, it can occur during the premenopausal andperimenopausal years. The use of oral contraceptives may also cause areduction in vaginal moisture in some women (W. Reginald, et al., Br. J.Obstet. Gynaecol. 96:1148-1152 (1989)). Postpartum vaginal dryness,independent of or as a result of lactation, can be a significantcomplaint (P. Wisniewski, et al., Am. J. Obstet. Gynecol. 165:1249-1254(1991)). Women undergoing chemotherapy or radiotherapy for malignantdiseases such as leukemia often experience vaginal dryness as a resultof treatment (M. Cust, et al., Br. Med. J. 299:1494-1497 (1989)). Manydisease states, such as systemic sclerosis and other systemic autoimmunedisorders (S. Bhadauria, et al., Am. J. Obstet. Gynecol. 172:580-587(1995)), Ehlers-Danlos syndrome (Y. Sorokin, et al., J. Reprod. Med.39:281-284 (1994)), diabetes mellitus (L. Sreebny, et al., Diabetes Care15:900-904 (1992)), and Sjögren's syndrome (D. Marchesoni, et al., Eur.J. Obstet. Gynecol. Reprod Biol. 63:49-53 (1995)) have decreased vaginalhydration and lubrication problems as significant disease-associatedsymptoms.

[0007] Current therapies for increasing vaginal moisture are:lubricating agents such as lubricating creams or jellies, topicalestrogen creams, and HRT (hormone replacement therapy). Lubricatingjellies provide short-lived and temporary relief, as these are aqueouspreparations containing no pharmacologically active agent. Topicalestrogen creams, if used on a regular basis, may be absorbed into thesystemic circulation. This can cause endometrial stimulation and canlead to endometrial hyperplasia and carcinoma (M. Whitehead, et al., N.Eng. J. Med. 305:1599-1605 (1981)). HRT is effective at relievingsymptoms of vaginal atrophy and hence vaginal dryness but has severalcontraindications and unwanted risks and side effects.

[0008] Another disease state characterized by improper hydration ofmucus secretions is acute and chronic sinusitis, or rhinosinusitis as itis currently referred to by otolaryngologists. Rhinosinusitis is adisease of the paranasal sinuses typically associated with retained,thick mucus secretions. It is this country's most common health-carecomplaint, affecting an estimated 31 million people (A. Moss and V.Parsons, National Center for Health Statistics, 1986: 66-7, DHHSPublication No. (PHS) 86-1588 (1985)).

[0009] Dry mouth is a condition in which the mucus membranes of theoropharyngeal cavity becomes dehydrated, leading to discomfort,difficulty swallowing and bad breath. Among the conditions generallyreferred to as “dry mouth” are, but not limited to: Sjogren's syndrome,chemotherapy and post radiation treatment, and pharmacologic sideeffects.

[0010] Thus, as a result of the ineffectiveness and risks of currenttherapies, medical researchers have sought to develop alternatives forthe treatment of dehydrated mucus membranes. Use of UDP and ADP for thepurpose of treating pulmonary disorders characterized by the retentionof lung mucus secretions is described in Patent Application (Boucher, R.C., Jr., et al., PCT/US98/17894). Applicants were motivated toinvestigate whether UDP and other P2Y₆ receptor agonists could stimulatehydration and mucin production on mucosal surfaces throughout the body,such as in the eye, female reproductive tract, sinonasal andoropharyngeal cavities.

[0011] Applicant has discovered that ion, mucin and fluid secretion canbe stimulated from mucous membrane tissues via P2Y₆ purinergicreceptor-mediated mechanisms. UDP and other purinergic receptoragonists, administered topically or systemically, provide a novel methodof treating disorders of mucosal hydration.

[0012] Applicant has also discovered that UDP and other diphosphateagonists of the P2Y₆ receptor also facilitate expectoration of deep lungmucus. Such expectoration is useful in a method of diagnosing lungdiseases whereby the expectorated deep lung mucus is analyzed forcytological abnormalities indicative of lung disease, such as lungcancer and tubeculosis.

SUMMARY OF THE INVENTION

[0013] A method of stimulating mucosal hydration in a subject in need ofsuch treatment is disclosed. The method of the present invention may beused to increase mucosal hydration for any reason, including, but notlimited to, treatment of dry eye disease, vaginal dryness,rhinosinusitis and dry mouth.

[0014] Dry eye disease is defined to include: keratoconjunctivitis sicca(KCS), age-related dry eye, Stevens-Johnson syndrome, Sjogren'ssyndrome, ocular cicatrical pemphigoid, blepharitis, corneal injury,infection, Riley-Day syndrome, congenital alacrima, nutritionaldisorders or deficiencies (including vitamin), pharmacologic sideeffects, eye stress and glandular and tissue destruction, environmentalexposure to smog, smoke, excessively dry air, airborne particulates,autoimmune and other immunodeficient disorders, and comatose patientsrendered unable to blink. The present invention may also be useful as anophthalmic wash or irrigation solution in conscious individuals, duringsurgery (e.g. lasik, radial keratotomy, cataract removal) or to maintaincomatose patients or those who cannot blink due to neuromuscularblockade, muscle or nerve damage, or loss of the eyelids. The compounduridine diphosphate (UDP) was found to be a potent stimulator of mucinsecretion in tissue preparations containing goblet cells. Furthermore,an in vivo example of mucin secretion in accordance with the inventionis conducted on an animal using impression cytology.

[0015] A method of stimulating cervical and vaginal secretions in asubject in need of such treatment is disclosed. The method of thepresent invention may be used to increase cervical and vaginalsecretions for any reason, including, but not limited to, treatment ofvaginal dryness. Vaginal dryness is associated with but not limited tomenopause, childbirth, breastfeeding, chemotherapy or radiotherapy,diabetes mellitus, Sjögren's syndrome, Ehlers-Danlos syndrome, systemicsclerosis and other systemic autoimmune diseases, hysterectomy,urogenital surgery, psychosomatic disorders, anxiety, psychosexualproblems, and pharmacological drug-related side effects.

[0016] Furthermore, because of their general ability to rehydratemucosal surfaces, the compounds of the present invention may also beuseful in the treatment of rhinosinusitis and dry mouth. Additionally,it is postulated that the compounds of the present invention could beuseful for the treatment of dry mouth.

[0017] The method of the present invention comprises administering aP2Y₆ receptor agonist: uridine 5′-diphosphate,P¹,P³-di(uridine-5′)triphosphate, cytidine 5′-diphosphate or adenosine5′-diphosphate or analogs thereof, in an amount effective to stimulatemucosal hydration.

[0018] Another aspect of the present invention is the use of uridine5′-diphosphate, P¹,P³-di(uridine-5′)triphosphate, cytidine5′-diphosphate or adenosine 5′-diphosphate or analogs thereof, for themanufacture of a medicament for carrying out a therapeutic method oftreatment as given above.

[0019] The present invention also discloses pharmaceutical compositionscomprising uridine 5′-diphosphate, P¹,P³-di(uridine-5′)triphosphate,cytidine 5′-diphosphate or adenosine 5′-diphosphate or analogs thereof,with a pharmaceutical carrier therefor.

[0020] The present invention also discloses a method of detecting lungdisease comprising administering uridine 5′-diphosphate, P¹,P³-di(uridine-5′)triphosphate, cytidine 5′-diphosphate or adenosine5′-diphosphate or analogs thereof, to at least one lung of an individualin order to facilitate the obtaining of a deep lung mucus, i.e., sputumsample, whereby the sputum sample is subjected to cytological,immunocytochemical, bacterial, or DNA analysis (e.g., via PCR) fordetecting cellular abnormalities in the lung epithelium. The lungdiseases detectable by this method include, but are not limited to lungcancer and tuberculosis. A preferred diphosphate compound for thisdiagnostic method is UDP (uridine 5′-diphosphate).

BRIEF DESCRIPTION OF THE FIGURES

[0021]FIG. 1 shows mucin secretion in normal human bronchial/trachealepithelia cells (Donor #8F0258) in response to different concentrationsof UDP.

DETAILED DESCRIPTION OF THE INVENTION

[0022] The method of the present invention may be used to increasemucosal hydration for any reason, including, but not limited to,treatment of dry eye disease, vaginal dryness, rhinosinusitis and drymouth. Dry eye disease is defined to include: keratoconjunctivitis sicca(KCS), age-related dry eye, Stevens-Johnson syndrome, Sjogren'ssyndrome, ocular cicatrical pemphigoid, blepharitis, corneal injury,infection, Riley-Day syndrome, congenital alacrima, nutritionaldisorders or deficiencies (including vitamin), pharmacologic sideeffects, eye stress and glandular and tissue destruction, environmentalexposure to smog, smoke, excessively dry air, airborne particulates,autoimmune and other immunodeficient disorders, and comatose patientsrendered unable to blink. The present invention may also be useful as anophthalmic wash or irrigation solution in conscious individuals, duringsurgery or to maintain comatose patients or those who cannot blink dueto muscle or nerve damage, neuromuscular blockade or loss of theeyelids.

[0023] Applicants have discovered that uridine 5′-diphosphate (UDP) andrelated compounds are potent agonists for purinergic receptors found incervical and vaginal mucus membrane preparations. The methods of thepresent invention are an improvement upon the current most commonly usedtreatments of vaginal dryness because UDP stimulates a patient's ownproduction and secretion of mucins as well as increasing the levels ofmucosal hydration, which serve to maintain the natural protective andlubricant characteristics of vaginal and cervical mucosa. The methods ofthe present invention may also be used exclusive of, or as an adjunctto, hormone replacement therapy (HRT) or estrogen replacement therapy(ERT).

[0024] Another disease state characterized by improper hydration ofmucus secretions is sinusitis. Sinusitis is a disease of the paranasalsinuses typically associated with retained, thick mucus secretions.

[0025] Dry mouth is a condition in which the mucus membranes of theoropharyngeal cavity becomes dehydrated, leading to discomfort,difficulty swallowing and bad breath. Among the conditions generallyreferred to as “dry mouth” are, but not limited to: Sjogren's syndrome,chemotherapy and post radiation treatment, and pharmacologic sideeffects.

[0026] Applicant has discovered that uridine 5′-diphosphate (UDP) is apotent agonist for purinergic receptors found in mucus membranepreparations. The method of the present invention is an improvement uponthe current most commonly used treatment of impaired mucosalhydration—artificial tears (i.e., saline solution) for dry eye; oilylubricants for vaginal drynesss; saline irrigation or decongestants forsinusitis; artificial saliva or Salagen for dry mouth. Because UDPstimulates a patient's own mucosal hydration mechanisms, it maintainsthe natural protective and lubricant characteristics. Furthermore, themethod of the present invention may be useful even where mucosal glandsare dysfunctional or absent.

[0027] The present invention also discloses a method of detecting lungdisease comprising administering uridine 5′-diphosphate,P¹,P³-di(uridine-5′)triphosphate, cytidine 5′-diphosphate or adenosine5′-diphosphate or analogs thereof, to at least one lung of an individualin order to facilitate the obtaining of a deep lung mucus, i.e., sputumsample, whereby the sputum sample is subjected to cytological,immunocytochemical, bacterial, or DNA analysis (e.g., via PCR) fordetecting cellular abnormalities in the lung epithelium. The lungdiseases detectable by this method include, but are not limited to lungcancer and tuberculosis. A preferred diphosphate compound for thisdiagnostic method is UDP (uridine 5′-diphosphate). Prior to analysis ofthe sputum sample, the sputum sample may first be digested with aliquefying agent, such as N-acetyl-L-cystein (NALC) and sodiumhydroxide.

[0028] The present invention provides a method of stimulating mucosalhydration in a mammal, including a human, in need thereof comprisingadministering an amount of a compound of Formulas I, II, III, or IV or apharmaceutically acceptable ester or salt thereof effective to increasesaid hydration.

[0029] UDP and its analogs are depicted in general Formula I:

[0030] wherein:

[0031] X₁ and X₂ are each independently either O⁻ or S⁻; preferably, X₁and X₂ are O⁻;

[0032] R₁ is O, imido, methylene or dihalomethylene (e.g.,dichloromethylene or difluoromethylene); preferably, R₁ is oxygen ordifluoromethylene;

[0033] R₂ is selected from the group consisting of H, halo, alkyl,substituted alkyl, optionally substituted alkenyl, optionallysubstituted alkynyl, alkoxyl, nitro and azido; preferably, R₂ is H;particularly preferred compounds of Formula I are uridine 5′-diphosphate(UDP) and uridine 5′-O-(2-thiodiphosphate) (UDPβS).

[0034] A dinucleotide is depicted by the general Formula II:

[0035] wherein:

[0036] X is oxygen, methylene, difluoromethylene, or imido;

[0037] B and B′ are each independently a purine residue or a pyrimidineresidue linked through the 9- or 1- position, respectively;

[0038] Z═OH or N₃;

[0039] Z′═OH or N₃;

[0040] Y═H or OH;

[0041] Y′═H or OH;

[0042] provided that when Z is N₃, Y is H or when Z′ is N₃, Y′ is H.

[0043] The furanose sugar is preferably in the β-configuration.

[0044] The furanose sugar is most preferably in the β-D-configuration.

[0045] Preferred compounds of Formula II are the compounds of FormulaIIa:

[0046] wherein:

[0047] X═O;

[0048] Z, Z′, Y, and Y′═OH;

[0049] B and B′ are defined in Formulas IIc and IId;

[0050] X═O;

[0051] Z, Z′, Y, and Y′═OH;

[0052] B=uracil;

[0053] B′ is defined in Formulas IIc and IId; or

[0054] X═O;

[0055] Z, Y, and Z¹═OH;

[0056] Y¹═H;

[0057] B=uracil;

[0058] B′ is defined in Formulas IIc and IId; or

[0059] X═O;

[0060] Z and Y═OH;

[0061] Z′═N₃;

[0062] Y′═H;

[0063] B=uracil;

[0064] B′=thymine; or

[0065] X═O;

[0066] Z and Z′═N₃;

[0067] Y and Y′═H;

[0068] B and B′=thymine; or

[0069] X═CH₂, CF₂, or NH;

[0070] Z, Z′, Y, and Y′═OH;

[0071] B and B′ are defined in Formulas IIc and IId.

[0072] Another preferred group of the compounds of Formula II are thecompounds of Formula IIb or the pharmaceutically acceptable saltsthereof:

[0073] wherein:

[0074] X is oxygen, methylene, difluoromethylene, or imido; and

[0075] B and B′ are each independently a purine residue, as in FormulaIIc, or a pyrimidine residue, as in Formula IId, linked through the 9-or 1- position, respectively. In the instance where B and B′ are uracil,attached at N-1 position to the ribosyl moiety, then the total of m+nmay equal 3 or 4 when X is oxygen. The ribosyl moieties are in the D-configuration, as shown, but may be L-, or D- and L-. The D-configuration is preferred.

[0076] wherein

[0077] R₁ is hydrogen, C₁₋₈alkyl, C₃₋₆ cycloalkyl, phenyl, or phenyloxy;wherein at least one hydrogen of said C₁₋₈alkyl, phenyl, phenyloxy, isoptionally substituted with a moiety selected from the group consistingof halogen, hydroxy, C₁₋₄alkoxy, C₁₋₄alkyl, C₆₋₁₀aryl, carboxy, cyano,nitro, sulfonamido, sulfonate, phosphate, sulfonic acid, amino, C₁₋₄alkylamino, di-C₁₋₄ alkylamino wherein said alkyl groups are optionallylinked to form a heterocycle, ω-A(alkyl)CONH(alkyl)-, andω-A(alkyl)NHCO(alkyl)-, wherein A is amino, mercapto, hydroxy orcarboxyl;

[0078] R₂ is O or is absent; or

[0079] R₁ and R₂ taken together form a 5-membered fused imidazole ringoptionally substituted on the 4- or 5- positions of the etheno moietywith C₁₋₄alkyl, phenyl or phenyloxy, wherein at least one hydrogen ofsaid C₁₋₄alkyl, phenyl, phenyloxy, is optionally substituted with amoiety selected from the group consisting of halogen, hydroxy,C₁₋₄alkoxy, C₁₋₄alkyl, C₆₋₁₀aryl, C₇₋₁₂arylalkyl, carboxy, cyano, nitro,sulfonamido, sulfonate, phosphate, sulfonic acid, amino, C₁₋₄alkylamino, and di-C₁₋₄ alkylamino wherein said dialkyl groups areoptionally linked to form a heterocycle; and

[0080] R₃ is hydrogen, NH₂, C₁₋₈alkyl, C₃₋₆ cycloalkyl, phenyl; orphenyloxy; wherein at least one hydrogen of said NH₂, C₁₋₈ alkyl,phenyl, or phenyloxy, is optionally substituted with a moiety selectedfrom the group consisting of halogen, hydroxy, C₁₋₄alkyl, C₆₋₁₀aryl,C₇₋₁₂arylalkyl, C₁₋₄alkoxy, C₇₋₁₂arylalkyloxy, C₁₋₄alkylthio,phenylthio, C₇₋₁₂arylalkylthio, carboxy, cyano, nitro, sulfonamido,sulfonate, phosphate, sulfonic acid, amino, C₁₋₄ alkylamino,phenylamino, C₇₋₁₂arylalkyamino, di-C₁₋₄ alkyl amino wherein saiddialkyl groups are optionally linked to form a heterocycle,ω-A(alkyl)CONH(alkyl)B-, and ω-A(alkyl)NHCO(alkyl)B-, wherein A and Bare independently amino, mercapto, hydroxy or carboxyl.

[0081] The substituted derivatives of adenine (Formula IIc) includeadenine 1-oxide; 1,N6-(4- or 5-substituted etheno) adenine;6-substituted adenine; or 8-substituted aminoadenine,[6-aminohexyl]carbamoylmethyl-adenine; and ω-acylated- amino(hydroxy,thiol and carboxy)alkyl(C₂₋₁₀)-adenine, wherein the acyl group is chosenfrom among, but not limited to, acetyl, trifluroroacetyl, benzoyl,substituted-benzoyl, etc., or the carboxylic moiety is present as itsester or amide derivative, for example, the ethyl or methyl ester or itsmethyl, ethyl or benzamido derivative.

[0082] Likewise, B or B¹ or both in Formula IIb may be a pyrimidine withthe general formula of Figure IId, linked through the 1-position:

[0083] wherein:

[0084] R₄ is hydrogen, hydroxy, mercapto, amino, cyano, C₇₋₁₂arylalkoxy,C₁₋₆ alkylthio, C₁₋₆ alkoxy, C₁₋₆ alkylamino or diC₁₋₄alkylamino,wherein the alkyl groups are optionally linked to form a heterocycle;

[0085] R₅ is hydrogen, acetyl, benzoyl, C₁₋₆ alkyl, phenyloxy, C₁₋₅alkanoyl, aroyl, or sulphonate;

[0086] R₆ is hydroxy, mercapto, C₁₋₄alkoxy, C₇₋₁₂arylalkoxy,C₁₋₆alkylthio, amino, S-phenyl, C₁₋₅ disubstituted amino, triazolyl,C₁₋₆alkylamino, or di-C₁₋₄alkylamino wherein said dialkyl groups areoptionally linked to form a heterocycle or linked to N³ to form asubstituted ring; or

[0087] R₅ and R₆ taken together form a 5-membered fused imidazole ringbetween positions 3 and 4 of the pyrimidine ring and form a3,N⁴-ethenocytosine derivative, wherein said etheno moiety is optionallysubstituted on the 4- or 5- positions with C₁₋₄ alkyl; phenyl; orphenyloxy; wherein at least one hydrogen of said C₁₋₄alkyl; phenyl orphenyloxy is optionally substituted with a moiety selected from thegroup consisting of halogen, hydroxy, C₁₋₄alkoxy, C₁₋₄alkyl, C₆₋₁₀aryl,C₇₋₁₂arylalkyl, carboxy, cyano, nitro, sulfonamido, sulfonate,phosphate, sulfonic acid, amino, C₁₋₄ alkylamino, and di- C₁₋₄alkylamino wherein said dialkyl groups are optionally linked to form aheterocycle;

[0088] R₇ is hydrogen, hydroxy, cyano, nitro, or C₂₋₈alkenyl; whereinsaid alkenyl moiety is optionally linked through an oxygen to form aring, wherein at least one hydrogen of said alkenyl moiety on the carbonadjacent to said oxygen is optionally substituted with C₁₋₆alkyl,phenyl, substituted C₂₋₈alkynyl, halogen, substituted C₁₋₄alkyl, CF₃,C₂₋₃ alkenyl, C₂₋₃ alkynyl, alkylamino, bromovinyl, ethyl propenoate, orpropenoic acid; or

[0089] R₆ and R₇ together form a 5 or 6-membered saturated orunsaturated ring bonded through N or O at R₆, such ring optionallycontains substituents that themselves contain functionalities; providedthat when R₈ is amino or substituted amino, R₇ is hydrogen; and

[0090] R₈ is hydrogen, amino or di-C₁₋₄alkylamino, C₁₋₄alkoxy,C₇-₁₂arylalkoxy, C₁₋₄alkylthio, C₇₋₁₂arylalkylthio,carboxamidomethyl,carboxymethyl, methoxy, methylthio, phenoxy or phenylthio.

[0091] In the general structure of Figure IId above, the dotted lines inthe 2- to 6-positions are intended to indicate the presence of single ordouble bonds in these positions; the relative positions of the double orsingle bonds being determined by whether the R₄, R₆, and R₇ substituentsare capable of keto-enol tautomerism.

[0092] In the general structures of Figure IIc and IId above, the acylgroups advantageously comprise alkanoyl or aroyl groups. The alkylgroups advantageously contain 1 to 8 carbon atoms, particularly 1 to 4carbon atoms optionally substituted by one or more appropriatesubstituents, as described below. The aryl groups including the arylmoieties of such groups as aryloxy are preferably phenyl groupsoptionally substituted by one or more appropriate substituents, asdescribed below. The above mentioned alkenyl and alkynyl groupsadvantageously contain 2 to 8 carbon atoms, particulary 2 to 6 carbonatoms, e.g., ethenyl or ethynyl, optionally substituted by one or moreappropriate substituents as described below. Appropriate substituents onthe above-mentioned alkyl, alkenyl, alkynyl, and aryl groups areadvantageously selected from halogen, hydroxy, C₁₋₄ alkoxy, C₁₋₄ alkyl,C₆₋₁₂ arylalkoxy, carboxy, cyano, nitro, sulfonamido, sulfonate,phosphate, sulfonic, amino, and substituted amino wherein the amino issingly or doubly substituted by a C₁₋₄ alkyl, and when doublysubstituted, the alkyl groups optionally being linked to form aheterocycle.

[0093] CDP and its analogs are depicted by general Formula III:

[0094] wherein:

[0095] R₁, X₁ and X₂ are defined as in Formula I;

[0096] R₅ and R₆ are H while R₇ is nothing and there is a double bondbetween N-3 and C-4 (cytosine), or

[0097] R₅, R₆ and R₇ taken together are —CH═CH—, forming a ring from N-3to N-4 with a double bond between N-4 and C-4 (3,N⁴-ethenocytosine)optionally substituted at the 4- or 5-position of the etheno ring.

[0098] ADP and its analogs are depicted by general Formula IV:

[0099] wherein:

[0100] R₁, X₁ and X₂ are defined as in Formula I;

[0101] R₃ and R₄ are H while R₂ is nothing and there is a double bondbetween N-1 and C-6 (adenine), or

[0102] R₃ and R₄ are H while R₂ is O and there is a double bond betweenN-1 and C-6 (adenine 1-oxide), or

[0103] R₃, R₄, and R₂ taken together are —CH═CH—, forming a ring fromN-6 to N-1 with a double bond between N-6 and C-6 (1,N6-ethenoadenine).

[0104] For simplicity, Formulas I, II, III, and IV herein illustrate theactive compounds in the naturally occurring D-configuration, but thepresent invention also encompasses compounds in the L-configuration, andmixtures of compounds in the D- and L-configurations, unless otherwisespecified. The naturally occurring D-configuration is preferred.

[0105] The compounds of the invention may be present in the form oftheir pharmaceutically acceptable salts, such as, but not limited to, analkali metal salt such as sodium or potassium; an alkaline earth metalsalt such as manganese, magnesium, or calcium; or an ammonium ortetraalkyl ammonium salt, i.e., NX₄ ⁺ (wherein X is C₁₋₄).Pharmaceutically acceptable salts are salts that retain the desiredbiological activity of the parent compound and do not impart undesiredtoxicological effects.

[0106] The compounds of the invention may also be present in the form ofprodrugs, typically comprising esters or amide moieties on theheterocyclic and furanosyl hydroxyls of the compound.

[0107] Another aspect of the present invention is a method of treating amammal with vaginal dryness arising from, but not limited to, menopause,childbirth, breastfeeding, chemotherapy or radiotherapy, diabetesmellitus, Sjögren's syndrome, Ehlers-Danlos syndrome, systemic sclerosisand other systemic autoimmune diseases, hysterectomy, urogenitalsurgery, psychosomatic disorders, anxiety, psychosexual problems, andpharmacological drug-related side effects.

[0108] The present invention further provides pharmaceuticalcompositions comprising a P2Y₆ receptor agonist selected from the groupconsisting of general Formula I, i.e., uridine 5′-diphosphate [UDP] andits analogs, general Formula II, i.e., P¹,P⁴-di(uridine-5′) triphosphate[U₂P₃] and its analogs, general Formula III, i.e., cytidine5′-diphosphate [CDP] and its analogs, and general Formula IV, i.e.,adenosine 5′-diphosphate [ADP] and its analogs, together with apharmaceutical carrier therefor.

[0109] The compounds disclosed herein may be administered to the mucosaof a patient by any suitable means, but are preferably administered by asolution, gel, suspension, cream, foam, pessary, or tablet containingthe active compound. Alternatively, the active compounds may byadministered by continuous release from a vaginal ring (P. Stumpf,Obstet. Gynecol. 75:9S (1990)) or an intrauterine device (K. Andersson,et al., Obstet. Gynecol. 79:963 (1992)).

[0110] The active compounds disclosed herein may be administered to theeyes of a patient by any suitable means, but are preferably administeredby administering a liquid or gel suspension of the active compound inthe form of drops, spray or gel. Alternatively, the active compounds maybe applied to the eye via liposomes. Further, the active compounds maybe infused onto the mucosal surface via a pump-catheter system. Anotherembodiment of the present invention involves the active compoundcontained within a continuous or selective-release device, for example,membranes such as, but not limited to, those employed in the Ocusert™System (Alza Corp., Palo Alto, Calif.). As an additional embodiment, theactive compounds can be contained within, carried by, or attached tocontact lenses which are placed on the eye. Another embodiment of thepresent invention involves the active compound contained within a swabor sponge which can be applied to the ocular surface. Another embodimentof the present invention involves the active compound contained within aliquid spray which can be applied to the ocular surface.

[0111] The topical solution, gel, jelly, ointment, cream, foam, pessary,or tablet contain the active compound in a physiologically compatiblevehicle, as those skilled in the art of topical delivery systemdevelopment can select using conventional criteria.

[0112] Solutions formulated for administration to the vagina are usuallyreferred to as irrigations. These are sterile solutions, prepared in amanner typical of sterile injections that are intended for prepared as asingle use sterile solution.

[0113] Gels or jellies may be produced using a suitable gelling agentincluding, but not limited to, gelatin, tragacanth, or a cellulosederivative and may include glycerol as a humectant, emollient, andpreservative.

[0114] Ointments are semi-solid preparations that consist of the activeingredient incorporated into a fatty, waxy, or synthetic base.

[0115] Examples of suitable creams include, but are not limited to,water-in-oil and oil-in-water emulsions. Water-in-oil creams may beformulated by using a suitable emulsifying agent with propertiessimilar, but not limited, to those of the fatty alcohols such as cetylalcohol or cetostearyl alcohol and to emulsifying wax. Oil-in-watercreams may be formulated using an emulsifying agent such as cetomacrogolemulsifying wax. Suitable properties include the ability to modify theviscosity of the emulsion and both physical and chemical stability overa wide range of pH. The water soluble or miscible cream base may containa preservative system and may also be buffered to maintain an acceptablephysiological pH.

[0116] Foam preparations may be formulated to be delivered from apressurized aerosol canister, via a suitable applicator, using inertpropellants. Suitable excipients for the formulation of the foam baseinclude, but are not limited to, propylene glycol, emulsifying wax,cetyl alcohol, and glyceryl stearate. Potential preservatives includemethylparaben and propylparaben.

[0117] Pessaries are solid unit-dose forms suitably shaped for insertioninto the vagina and may either be composed of a base that melts at bodytemperature or which dissolves when in contact with mucous secretions.Examples of suitable bases include, but are not limited to, theobromaoil, synthetic fat bases (e.g. Witepsol), polyethylene glycols(macrogols), and glycerol suppository basis.

[0118] Tablets are composed of the active ingredient contained within asolid dosage form base which may include, but not be limited to,excipients such as lactose, microcrystalline cellulose, corn starch,magnesium stearate, silicon dioxide, and hydroxypropyl methylcellulose.

[0119] An alternative method of administering the compounds of thepresent invention is intraoperative instillation during surgicalprocedures involving mucosal surfaces of the eye, sinuses, mouth andfemale reproductive tract.

[0120] In addition to the topical method of administration describedabove, there are various methods of administering the compounds of thepresent invention systemically. One such means would involve an aerosolsuspension of respirable particles comprised of the active compound,which the subject inhales. The active compound would be absorbed intothe bloodstream via the lungs and contact the cervical and/or vaginaltissues in a pharmaceutically effective amount. The respirable particlesmay be liquid or solid, with a particle size sufficiently small to passthrough the mouth and larynx upon inhalation; in general, particlesranging from about 1 to 10 microns, but more preferably 1-5 microns, insize are considered respirable.

[0121] Another means of systemically administering the active compoundsto the mucosal tissues of the subject would involve administering aliquid/liquid suspension in the form of nasal drops of a liquidformulation, or a nasal spray of respirable particles which the subjectinhales. Liquid pharmaceutical compositions of the active compound forproducing a nasal spray or nasal drops may be prepared by combining theactive compound with a suitable vehicle, such as sterile pyrogen freewater or sterile saline by techniques known to those skilled in the art.

[0122] Other means of systemic administration of the active compoundwould involve oral administration, in which pharmaceutical compositionscontaining compounds of Formulas I, II, III, or IV are in the form oftablets, lozenges, aqueous or oily suspensions, dispersible powders orgranules, emulsion, hard or soft capsules, or syrups or elixirs.Compositions intended for oral use may be prepared according to anymethod known to the art for the manufacture of pharmaceuticalcompositions and such compositions may contain one or more agentsselected from the group consisting of sweetening agents, flavoringagents, coloring agents, and preserving agents in order to providepharmaceutically elegant and palatable preparations. Tablets contain theactive ingredient in admixture with nontoxic pharmaceutically acceptableexcipients which are suitable for the manufacture of tablets. Theseexcipients may be, for example, inert diluents, such as calciumcarbonate, sodium carbonate, lactose, calcium phosphate, or sodiumphosphate; granulating and disintegrating agents, for example, cornstarch or alginic acid; binding agents, for example, starch, gelatin, oracacia; and lubricating agents, for example magnesium stearate, stearicacid, or talc. The tablets may be uncoated or they may be coated byknown techniques to delay disintegration and absorption in thegastrointestinal tract and thereby provide a sustained action over alonger period. For example, a time delay material such as glycerylmonostearate or glyceryl distearate may be employed. Formulations fororal use may also be presented as hard gelatin capsules wherein theactive ingredient is mixed with an inert solid diluent, for example,calcium carbonate, calcium phosphate, or kaolin, or as soft gelatincapsules wherein the active ingredient is mixed with water or an oilmedium, for example, peanut oil, liquid paraffin, or olive oil.

[0123] Additional means of systemic administration of the activecompound to the cervical and vaginal tissues of the subject wouldinvolve a suppository form of the active compound, such that atherapeutically effective amount of the compound reaches the mucosaltissues via systemic absorption and circulation.

[0124] For the above mentioned indications, the quantity of the activecompound included in the pharmaceutical composition is an amountsufficient to achieve concentrations of the active compound on themucosa of the subject of from about 10⁻⁷ to about 10⁻¹ Moles/liter, andmore preferably from about 10⁻⁶ to about 10⁻¹ Moles/liter.

[0125] Depending upon the solubility of the particular formulation ofactive compound administered, the daily dose to promote tear secretionmay be divided among one or several unit dose administrations. The totaldaily dose for UDP (for example) may range from 0.2 mg to 300 mg,depending upon the age and condition of the subject. A currentlypreferred unit dose for UDP is about 0.1 to 50 milligrams given at aregimen of 2 to 6 administrations per day.

[0126] Depending on the solubility of the particular formulation ofactive compound administered, the daily dose to promote mucous membranemucin production and/or hydration may be divided among one or severalunit dose administrations. The total daily dose for UDP (for example)may range from 1 to 1000 milligrams, depending upon the age and state ofthe subject, given at a regimen of up to four times per day or on an asneeded basis to address acute exacerbations.

[0127] Some compounds of Formulas I, II, III, and IV can be made bymethods which are well known to those skilled in the art and inaccordance with known procedures (P. Zamecnik, et al., Proc. Natl Acad.Sci. USA 89:838-842 (1981); K. Ng et al., Nucleic Acids Res.15:3572-3580 (1977); K. M. Jacobus, et al., U.S. Pat. No. 5,789,391 andW. Pendergast, et al., International Patent No. WO98/34942; Boucher, RC, Jr. et al., PCT/US98/17894) which are incorporated herein byreference; some are commercially available, for example, from SigmaChemical Company, PO Box 14508, St. Louis, Mo. 63178. The syntheticmethods of U.S. Pat. No. 5,789,391 and International Patent No.WO98/34942 are incorporated herein by reference.

Example 1 UDP Stimulates Mucin Release From Human Mucosal EpithelialCell Culture

[0128] The following is an example showing mucin release in response toUDP in an in vitro cell culture system containing mucosal epithelial andgoblet cells. Mucins, a family of high-molecular-weight glycoproteinssecreted by goblet and non-goblet epithelial cells of mucus membranes,are responsible for mucosal hydration. The objective of this study wasto demonstrate the effect of UDP on mucin release in Normal HumanBronchial/Tracheal Epithelial cells (NHTBE).

[0129] NHTBE cells were obtained from a commercial source (Clonetics;CC-2540). The cells are donor specific (non-smoker) and shipped ascryopreserved primary cultures. Bronchial/Epithelial Growth Media (BEGM;Clonetics; CC-3170) was used for expansion. The BEGM media supplied wasmodified as follows: The level of EGF (Intergen; #4110-80) was increasedto a final concentration of 25 ng/ml; the level of Bovine PituitaryExtract (BPE, Clonetics, CC 4009) was increased to 0.13 mg/ml; the levelof retinoic acid (Sigma; R-2625) was increased to a final concentrationof 5×10-8 M. The cells for the expansion phase were seeded at 500cells/cm². Media was changed after 24 hours and every 48 hoursthereafter until cells are 75% confluent (5-14 days). It was importantnot to exceed 75% confluency. Cells were dissociated with trypsin. Oncecollected, cells were stored in liquid nitrogen at passage-2 [Freezingsolution: 80% media (BEGM), 10% FBS, 10% sterile DMSO].

[0130] NHTBE cells must undergo a program of differentiation in order toachieve a mucin-secreting phenotype. This was achieved by culturingcells on porous membrane cell culture inserts (Transwell™; 12 mm, 0.45μm pore size, Corning-Costar #3460) ultimately under conditions of anair/liquid interface. The insert membrane was first coated with rat tailcollagen (Collaborative Biomedical Products #40236) as follows: 235 μlof coating solution(50 μg/ml of rat tail collagen dissolved in 0.02Nacetic acid) was added to apical surface of insert. The insert wasallowed to dry at room temperature for one hour in tissue culture hoodand then washed once with 500 μl of PBS containing nystatin (20,000units/liter sigma;N-1638) and gentamicin (100 μg/ml). Next, the 500 μlPBS was aspirated and the inserts equilibrated with BEGM media in theapical and basal compartments; incubated for one hour. Frozen NHTBEcells were thawed rapidly and seeded on the collagen-coated inserts at20,000 cells/cm². Media was changed after 24 hours and every 48 hoursthereafter until the cells were 60-75% confluent (5-7 days in culture).At this point, the cultures were switched to air-liquid interface byremoving the media bathing the apical aspect of the cells. The mediabathing the basolateral aspect of the culture must be changed dailyuntil the time of experiment. The composition of this ALI media was asfollows: A 1:1 mixture of Bronchial epithelial cell growth medium:Dulbecco's Modified Eagles medium with high glucose (BEGM: DMEM-H),containing final concentration of 0.5 ng/ml hrEGF, 0.5 μ/mlhydrocortisone, 5 μg/ml insulin, 10 μg/ml transferrin, 0.5 μg/mlepinephrine, 6.5 ng/ml triiodothyronine, 50 μg/ml gentamicin, 50 ng/mlamphotericin-B (all supplied by Clonetics as SingleQuots), 8 mg/ml BPE,5×10⁻⁸ M all-trans retinoic acid (Sigma #R-2625), 1.5 μg/ml BSA(Intergen, #3310-80), and 20 U/ml nystatin (Sigma #N-1638). SingleQuotscan be purchased separately from Clonetics (#CC-4175). As has beenreported by Gray, et al., Am J. Resp. Cell Molec. Biol. 14(1):104-112(1996), mucin phenotype was seen by 14 days in culture (7 days ofair/liquid interface), and cilia were apparent by 21 days in culture (14days of air/liquid interface).

[0131] Ascites fluid containing the anti-mucin antibody designated 17Q2IgG was kindly provided by Professor Reen Wu; (Department of InternalMedicine, University of California at Davis). 17Q2 antibody was purifiedwith a Protein G column (Pierce #44441). Alkaline phosphatase wasconjugated to 17Q2 antibody using the EZ-Link Malemide AcitivatedAlkaline Phosphatase kit (Pierce #31486).

[0132] UDP (10⁻⁸˜10⁻⁴) was added to the apical aspect of thedifferentiated cultures At the end of the incubation period (two hours)the stimulation media was removed and stored at −80° C. until analysisfor mucin content. Mucin was quantitated by ELISA as described above.

[0133] UDP stimulated the release of mucin glycoproteins fromdifferentiated cultures of human airway epithelium (FIG. 1) in aconcentration-dependent manner. The EC₅₀ was calculated to be 5.55×10⁻⁷M.

Example 2 Stimulation of Mucin Release from Mucosal Goblet Cells

[0134] The following is an example of a method for measuring the effectsof UDP and analogs in vivo on mucin secretion from mucous membranesusing impression cytology. Impression cytology is a technique used tostain and identify mucin-containing goblet cells (Rolando, M., et al.,Adv. Exp. Med. Bio. 350:249 (1994)).

[0135] UDP or saline solution is applied to the mucosal surface andimpression cytology is performed 5, 15, 30 and 60 minutes afterapplication of solution. The specimens are stained with periodic acidand Schiff's reagent (AB-PAS), and the area of PAS staining is analyzedby compouter software (Winroof or BioQuant). A decrease in the area ofAB-PAS staining compared to saline control indicates that UDP stimulatesmucin secretion from goblet cells of mucus membranes.

[0136] The present invention is concerned primarily with the treatmentof human subjects, but may also be employed for the treatment of othermammalian subjects, such as dogs and cats, for veterinary purposes.

[0137] The topical solution containing the active compound may alsocontain a physiologically compatible vehicle, as those skilled in theophthalmic art can select using conventional criteria. The vehicles maybe selected from the known ophthalmic vehicles which include, but arenot limited to, saline solution, water polyethers such as polyethyleneglycol, polyvinyls such as polyvinyl alcohol and povidone, cellulosederivatives such as methylcellulose and hydroxypropyl methylcellulose,petroleum derivatives such as mineral oil and white petrolatum, animalfats such as lanolin, polymers of acrylic acid such ascarboxypolymethylene gel, vegetable fats such as peanut oil andpolysaccharides such as dextrans, and glycosaminoglycans such as sodiumhyaluronate and salts such as sodium chloride and potassium chloride.

[0138] An alternative method of administering the compounds of thepresent invention is intraoperative instillation during surgicalprocedures involving mucosal surfaces of the eye, sinuses, mouth andfemale reproductive tract.

[0139] The invention and the manner and process of making and using itare now described in such full, clear, concise and exact terms as toenable any person skilled in the art to which it pertains, to make anduse the same. It is to be understood that the foregoing describespreferred embodiments of the present invention and that modificationsmay be made therein without departing from the spirit or scope of thepresent invention as set forth in the claims. To particularly point outand distinctly claim the subject matter regarded as invention, thefollowing claims conclude this specification.

What is claimed is:
 1. A method of stimulating mucosal hydration in atissue other than the lung of a mammal in need thereof, comprisingadministering an effective hydration increasing amount of a compound ofFormula I, II, III, or IV:

wherein: X₁ and X₂ are each independently either O⁻ or S⁻; R₁ is O,imido, methylene or dihalomethylene; R₂ is selected from the groupconsisting of: H, halo, alkyl, substituted alkyl, alkenyl andsubstituted alkenyl, alkynyl and substituted alkynyl, alkoxyl, nitro,and azido;

wherein: X is oxygen, methylene, difluoromethylene, or imido; B and B′are each independently a purine residue or a pyrimidine residue asdepicted in Formula IIc or IId, linked through the 9- or 1-position,respectively; Z═OH or N₃; Z′═OH or N₃; Y═H or OH; Y′═H or OH; providedthat when Z is N₃, Y is H or when Z′ is N₃, Y′ is H;

wherein R₁ is hydrogen, C₁₋₈alkyl, C₃₋₆ cycloalkyl, phenyl, orphenyloxy; wherein at least one hydrogen of said C₁₋₈alkyl, phenyl,phenyloxy, is optionally substituted with a moiety selected from thegroup consisting of: halogen, hydroxy, C₁₋₄alkoxy, C₁₋₄alkyl, C₆₋₁₀aryl,carboxy, cyano, nitro, sulfonamido, sulfonate, phosphate, sulfonic acid,amino, C₁₋₄ alkylamino, and di-C₁₋₄ alkylamino wherein said alkyl groupsare optionally linked to form a heterocycle, ω-A(alkyl)CONH(alkyl)-, andω-A(alkyl)NHCO(alkyl)-, wherein A is amino, mercapto, hydroxy, orcarboxyl; R₂ is O or is absent; or R₁ and R₂ taken together form a5-membered fused imidazole ring optionally substituted on the 4- or 5-positions of the etheno moiety with C₁₋₄alkyl, phenyl or phenyloxy,wherein at least one hydrogen of said C₁₋₄alkyl, phenyl or phenyloxy, isoptionally substituted with a moiety selected from the group consistingof: halogen, hydroxy, C₁₋₄alkoxy, C₁₋₄alkyl, C₆₋₁₀aryl, C₇₋₁₂arylalkyl,carboxy, cyano, nitro, sulfonamido, sulfonate, phosphate, sulfonic acid,amino, C₁₋₄ alkylamino, and di-C₁₋₄ alkylamino wherein said dialkylgroups are optionally linked to form a heterocycle; and R₃ is hydrogen,NH₂, C₁₋₈alkyl, C₃₋₆ cycloalkyl, phenyl, or phenyloxy, wherein at leastone hydrogen of said NH₂, C₁₋₈ alkyl, phenyl, or phenyloxy, isoptionally substituted with a moiety selected from the group consistingof: halogen, hydroxy, C₁₋₄alkyl, C₆₋₁₀aryl, C₇₋₁₂arylalkyl, C₁₋₄alkoxy,C₇₋₁₂arylalkyloxy, C₁₋₄alkylthio, phenylthio, C₇-₁₂arylalkylthio,carboxy, cyano, nitro, sulfonamido, sulfonate, phosphate, sulfonic acid,amino, C₁₋₁₄ alkylamino, phenylamino, C₇₋₁₂arylalkyamino, di-C₁₋₄ alkylamino, wherein said di-C₁₋₄ alkyl groups are optionally linked to form aheterocycle, ω-A(alkyl)CONH(alkyl)B-, and ω-A(alkyl)NHCO(alkyl)B-,wherein A and B are independently amino, mercapto, hydroxy, or carboxyl;

wherein: R₄ is hydrogen, hydroxy, mercapto, amino, cyano,C₇₋₁₂arylalkoxy, C₁₋₆ alkylthio, C₁₋₆ alkoxy, C₁₋₆ alkylamino, ordiC₁₋₄alkylamino, wherein the alkyl groups are optionally linked to forma heterocycle; R₅ is hydrogen, acetyl, benzoyl, C₁₋₆ alkyl, phenyloxy,C₁₋₅ alkanoyl, aroyl, or sulphonate; R₆ is hydroxy, mercapto,C₁₋₄alkoxy, C₇₋₁₂arylalkoxy, C₁₋₆alkylthio, amino, S-phenyl, C₁₋₅disubstituted amino, triazolyl, C₁₋₆alkylamino, or di-C₁₋₄alkylaminowherein said dialkyl groups are optionally linked to form a heterocycleor linked to N³ to form a substituted ring; or R₅ and R₆ taken togetherform a 5-membered fused imidazole ring between positions 3 and 4 of thepyrimidine ring and form a 3,N⁴-ethenocytosine derivative, wherein saidetheno moiety is optionally substituted on the 4- or 5-positions withC₁₋₄ alkyl, phenyl or phenyloxy, wherein at least one hydrogen of saidC₁₋₄alkyl, phenyl or phenyloxy is optionally substituted with a moietyselected from the group consisting of: halogen, hydroxy, C₁₋₄alkoxy,C₁₋₄alkyl, C₆₋₁₀aryl, C₇₋₁₂arylalkyl, carboxy, cyano, nitro,sulfonamido, sulfonate, phosphate, sulfonic acid, amino, C₁₋₄alkylamino, and di- C₁₋₄ alkylamino, wherein said dialkyl groups areoptionally linked to form a heterocycle; R₇ is hydrogen, hydroxy, cyano,nitro, or C₂₋₈alkenyl; wherein said alkenyl moiety is optionally linkedthrough an oxygen to form a ring, wherein at least one hydrogen of saidalkenyl moiety on the carbon adjacent to said oxygen is optionallysubstituted with C₁₋₆alkyl, phenyl, substituted C₂₋₈alkynyl, halogen,substituted C₁₋₄alkyl, CF₃, C₂₋₃ alkenyl, C₂₋₃ alkynyl, allylamino,bromovinyl, ethyl propenoate, or propenoic acid; or R₆ and R₇ togetherform a 5- or 6-membered saturated or unsaturated ring bonded through Nor O at R₆, such ring optionally contains substituents that themselvescontain functionalities; provided that when R₈ is amino or substitutedamino, R₇ is hydrogen; and R₈ is hydrogen, amino or di-C₁₋₄alkylamino,C₁₋₄alkoxy, C₇₋₁₂arylalkoxy, C₁₋₄alkylthio,C₇₋₁₂arylalkylthio,carboxamidomethyl, carboxymethyl, methoxy,methylthio, phenoxy, or phenylthio;

wherein: R₁, X₁ and X₂ are defined as in Formula I; R₅ and R₆ are Hwhile R₇ is nothing and there is a double bond between N-3 and C-4, orR₅, R₆ and R₇ taken together are —CH═CH—, forming a ring from N-3 to N-4with a double bond between N-4 and C-4 optionally substituted at the 4-or 5-position of the etheno ring;

wherein: R₁, X₁ and X₂ are defined as in Formula I; R₃ and R₄ are Hwhile R₂ is nothing and there is a double bond between N-1 and C-6, orR₃ and R₄ are H while R₂ is O and there is a double bond between N-1 andC-6, or R₃, R₄, and R₂ taken together are —CH═CH—, forming a ring fromN-6 to N-1 with a double bond between N-6 and C-6.
 2. A method ofstimulating mucosal hydration in a tissue selected from the groupconsisting of eye, vagina, sinonasal cavities and mouth of a mammal inneed thereof, comprising administering an effective hydration increasingamount of a compound of Formula I, II, III, or IV:

wherein: X₁ and X₂ are each independently either O⁻ or S⁻; R₁ is Oimido, methylene or dihalomethylene; R₂ is selected from the groupconsisting of: H, halo, alkyl, substituted alkyl, alkenyl andsubstituted alkenyl, alkynyl and substituted alkynyl, alkoxyl, nitro,and azido;

wherein: X is oxygen, methylene, difluoromethylene, or imido; B and B′are each independently a purine residue or a pyrimidine residue asdepicted in Formula IIc or IId, linked through the 9- or 1-position,respectively; Z═OH or N₃; Z′═OH or N₃; Y═H or OH; Y′═H or OH; providedthat when Z is N₃, Y is H or when Z′ is N₃, Y′ is H;

wherein R₁ is hydrogen, C₁₋₈alkyl, C₃₋₆ cycloalkyl, phenyl, orphenyloxy; wherein at least one hydrogen of said C₁₋₈alkyl, phenyl,phenyloxy, is optionally substituted with a moiety selected from thegroup consisting of: halogen, hydroxy, C₁₋₄alkoxy, C₁₋₄alkyl, C₆₋₁₀aryl,carboxy, cyano, nitro, sulfonamido, sulfonate, phosphate, sulfonic acid,amino, C₁₋₄ alkylamino, and di-C₁₋₄ alkylamino wherein said alkyl groupsare optionally linked to form a heterocycle, ω-A(alkyl)CONH(alkyl)-, andω-A(alkyl)NHCO(alkyl)-, wherein A is amino, mercapto, hydroxy, orcarboxyl; R₂ is O or is absent; or R₁ and R₂ taken together form a5-membered fused imidazole ring optionally substituted on the 4- or 5-positions of the etheno moiety with C₁₋₄alkyl, phenyl or phenyloxy,wherein at least one hydrogen of said C₁₋₄alkyl, phenyl or phenyloxy, isoptionally substituted with a moiety selected from the group consistingof: halogen, hydroxy, C₁₋₄alkoxy, C₁₋₄alkyl, C₆₋₁₀aryl, C₇₋₁₂arylalkyl,carboxy, cyano, nitro, sulfonamido, sulfonate, phosphate, sulfonic acid,amino, C₁₋₄ alkylamino, and di-C₁₋₄ alkylamino wherein said dialkylgroups are optionally linked to form a heterocycle; and R₃ is hydrogen,NH₂, C₁₋₈alkyl, C₃₋₆ cycloalkyl, phenyl, or phenyloxy, wherein at leastone hydrogen of said NH₂, C₁₋₈ alkyl, phenyl, or phenyloxy, isoptionally substituted with a moiety selected from the group consistingof: halogen, hydroxy, C₁₋₄alkyl, C₆₋₁₀aryl, C₇₋₁₂arylalkyl, C₁₋₄alkoxy,C₇₋₁₂arylalkyloxy, C₁₋₄alkylthio, phenylthio, C₇₋₁₂arylalkylthio,carboxy, cyano, nitro, sulfonamido, sulfonate, phosphate, sulfonic acid,amino, C₁₋₄ alkylamino, phenylamino, C₇₋₁₂arylalkyamino, and di-C₁₋₄alkyl amino, wherein said di-C₁₋₄ alkyl groups are optionally linked toform a heterocycle, ω-A(alkyl)CONH(alkyl)B -, andω-A(alkyl)NHCO(alkyl)B-, wherein A and B are independently amino,mercapto, hydroxy, or carboxyl;

wherein: R₄ is hydrogen, hydroxy, mercapto, amino, cyano,C₇₋₁₂arylalkoxy, C₁₋₆ alkylthio, C₁₋₆ alkoxy, C₁₋₆ alkylamino, ordiC₁₋₄alkylamino, wherein the alkyl groups are optionally linked to forma heterocycle; R₅ is hydrogen, acetyl, benzoyl, C₁₋₆ alkyl, phenyloxy,C₁₋₅ alkanoyl, aroyl, or sulphonate; R₆ is hydroxy, mercapto,C₁₋₄alkoxy, C₇₋₁₂arylalkoxy, C₁₋₆alkylthio, amino, S-phenyl, C₁₋₅disubstituted amino, triazolyl, C₁₋₆alkylamino, or di-C₁₋₄alkylaminowherein said dialkyl groups are optionally linked to form a heterocycleor linked to N³ to form a substituted ring; or R₅ and R₆ taken togetherform a 5-membered fused imidazole ring between positions 3 and 4 of thepyrimidine ring and form a 3,N⁴-ethenocytosine derivative, wherein saidetheno moiety is optionally substituted on the 4- or 5-positions withC₁₋₄ alkyl, phenyl or phenyloxy, wherein at least one hydrogen of saidC₁₋₄alkyl, phenyl or phenyloxy is optionally substituted with a moietyselected from the group consisting of: halogen, hydroxy, C₁₋₄alkoxy,C₁₋₄alkyl, C₆₋₁₀aryl, C₇₋₁₂arylalkyl, carboxy, cyano, nitro,sulfonamido, sulfonate, phosphate, sulfonic acid, amino, C₁₋₄alkylamino, and di- C₁₋₄ alkylamino, wherein said dialkyl groups areoptionally linked to form a heterocycle; R₇ is hydrogen, hydroxy, cyano,nitro, or C₂₋₈alkenyl; wherein said alkenyl moiety is optionally linkedthrough an oxygen to form a ring, wherein at least one hydrogen of saidalkenyl moiety on the carbon adjacent to said oxygen is optionallysubstituted with C₁₋₆alkyl, phenyl, substituted C₂-₈alkynyl, halogen,substituted C₁₋₄alkyl, CF₃, C₂₋₃ alkenyl, C₂₋₃ alkynyl, allylamino,bromovinyl, ethyl propenoate, or propenoic acid; or R₆ and R₇ togetherform a 5- or 6-membered saturated or unsaturated ring bonded through Nor O at R₆, such ring optionally contains substituents that themselvescontain functionalities; provided that when R₈ is amino or substitutedamino, R₇ is hydrogen; and R₈ is hydrogen, amino or di-C₁₋₄alkylamino,C₁₋₄alkoxy, C₇₋₁₂arylalkoxy, C₁₋₁₄alkylthio,C₇₋₁₂arylalkylthio,carboxamidomethyl, carboxymethyl, methoxy,methylthio, phenoxy, or phenylthio;

wherein: R₁, X₁ and X₂ are defined as in Formula I; R₅ and R₆ are Hwhile R₇ is nothing and there is a double bond between N-3 and C-4, orR₅, R₆ and R₇ taken together are —CH═CH—, forming a ring from N-3 to N-4with a double bond between N-4 and C-4 optionally substituted at the 4-or 5-position of the etheno ring;

wherein: R₁, X₁ and X₂ are defined as in Formula I; R₃ and R₄ are Hwhile R₂ is nothing and there is a double bond between N-1and C-6, or R₃and R₄ are H while R₂ is O and there is a double bond between N-1 andC-6, or R₃, R₄, and R₂ taken together are —CH═CH—, forming a ring fromN-6 to N-1 with a double bond between N-6 and C-6.
 3. The method ofclaim 2, wherein the effective hydration increasing amount of compoundof Formula I, II, III, or IV is administered to the eyes to treat dryeye disease.
 4. The method of claim 2, wherein the effective hydrationincreasing amount of compound of Formula I, II, III, or IV isadministered to the eyes to treat corneal injury.
 5. The method of claim2, wherein the effective hydration increasing amount of compound ofFormula I, II, III, or IV is administered to the sinuses to treatrhinosinusitis.
 6. The method of claim 2, wherein the effectivehydration increasing amount of compound of Formula I, II, III, or IV isadministered to the mouth to treat dry mouth disease.
 7. The method ofclaim 2, wherein the effective hydration increasing amount of compoundof Formula I, II, III, or IV is administered to the vagina to treatvaginal dryness.
 8. The method of claim 3, wherein said compound is UDP.9. A method for detecting a lung disease comprising the steps of:administering to at least one lung of a subject an expectorationincreasing amount of a compound of Formula I, II, III, or IV as depictedin claim 1 to facilitate the expectoration of sputum; obtaining a deeplung mucus from a sputum sample; subjecting said deep lung mucus to ananalysis; and detecting cellular abnormalities in the lung ephitheliumn.10. The method of claim 9, wherein the lung disease is lung cancer ortuberculosis.
 11. The method of claim 9, wherein the analysis comprisescytological analysis, immunocytochemical analysis, bacterial analysisand DNA analysis.
 12. The method of claim 9, wherein said compound isUDP.